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1.
Lancet Infect Dis ; 24(7): e453-e462, 2024 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-38340735

RESUMEN

SARS-CoV-2 causes persistent infections in a subset of individuals, which is a major clinical and public health problem that should be prioritised for further investigation for several reasons. First, persistent SARS-CoV-2 infection often goes unrecognised, and therefore might affect a substantial number of people, particularly immunocompromised individuals. Second, the formation of tissue reservoirs (including in non-respiratory tissues) might underlie the pathophysiology of the persistent SARS-CoV-2 infection and require new strategies for diagnosis and treatment. Finally, persistent SARS-CoV-2 replication, particularly in the setting of suboptimal immune responses, is a possible source of new, divergent virus variants that escape pre-existing immunity on the individual and population levels. Defining optimal diagnostic and treatment strategies for patients with persistent virus replication and monitoring viral evolution are therefore urgent medical and public health priorities.


Asunto(s)
COVID-19 , SARS-CoV-2 , Humanos , COVID-19/epidemiología , Replicación Viral , Huésped Inmunocomprometido
2.
PLoS Pathog ; 15(1): e1007518, 2019 01.
Artículo en Inglés | MEDLINE | ID: mdl-30673779

RESUMEN

Translation can initiate at alternate, non-canonical start codons in response to stressful stimuli in mammalian cells. Recent studies suggest that viral infection and anti-viral responses alter sites of translation initiation, and in some cases, lead to production of novel immune epitopes. Here we systematically investigate the extent and impact of alternate translation initiation in cells infected with influenza virus. We perform evolutionary analyses that suggest selection against non-canonical initiation at CUG codons in influenza virus lineages that have adapted to mammalian hosts. We then use ribosome profiling with the initiation inhibitor lactimidomycin to experimentally delineate translation initiation sites in a human lung epithelial cell line infected with influenza virus. We identify several candidate sites of alternate initiation in influenza mRNAs, all of which occur at AUG codons that are downstream of canonical initiation codons. One of these candidate downstream start sites truncates 14 amino acids from the N-terminus of the N1 neuraminidase protein, resulting in loss of its cytoplasmic tail and a portion of the transmembrane domain. This truncated neuraminidase protein is expressed on the cell surface during influenza virus infection, is enzymatically active, and is conserved in most N1 viral lineages. We do not detect globally higher levels of alternate translation initiation on host transcripts upon influenza infection or during the anti-viral response, but the subset of host transcripts induced by the anti-viral response is enriched for alternate initiation sites. Together, our results systematically map the landscape of translation initiation during influenza virus infection, and shed light on the evolutionary forces shaping this landscape.


Asunto(s)
Infecciones por Orthomyxoviridae/genética , Orthomyxoviridae/genética , Iniciación de la Cadena Peptídica Traduccional/genética , Animales , Aves/genética , Codón/genética , Codón Iniciador/genética , Codón Iniciador/metabolismo , Humanos , Subtipo H5N1 del Virus de la Influenza A/genética , Gripe Aviar/genética , Gripe Humana/genética , Orthomyxoviridae/patogenicidad , Infecciones por Orthomyxoviridae/metabolismo , Biosíntesis de Proteínas , Procesamiento Proteico-Postraduccional/genética , Proteínas/metabolismo , Proteómica/métodos , ARN Mensajero/metabolismo , Ribosomas/metabolismo , Homología de Secuencia de Aminoácido , Porcinos/virología , Transcriptoma/genética
3.
J Virol ; 89(22): 11275-83, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26311880

RESUMEN

UNLABELLED: Numerous experimental studies have demonstrated that CD8(+) T cells contribute to immunity against influenza by limiting viral replication. It is therefore surprising that rigorous statistical tests have failed to find evidence of positive selection in the epitopes targeted by CD8(+) T cells. Here we use a novel computational approach to test for selection in CD8(+) T-cell epitopes. We define all epitopes in the nucleoprotein (NP) and matrix protein (M1) with experimentally identified human CD8(+) T-cell responses and then compare the evolution of these epitopes in parallel lineages of human and swine influenza viruses that have been diverging since roughly 1918. We find a significant enrichment of substitutions that alter human CD8(+) T-cell epitopes in NP of human versus swine influenza virus, consistent with the idea that these epitopes are under positive selection. Furthermore, we show that epitope-altering substitutions in human influenza virus NP are enriched on the trunk versus the branches of the phylogenetic tree, indicating that viruses that acquire these mutations have a selective advantage. However, even in human influenza virus NP, sites in T-cell epitopes evolve more slowly than do nonepitope sites, presumably because these epitopes are under stronger inherent functional constraint. Overall, our work demonstrates that there is clear selection from CD8(+) T cells in human influenza virus NP and illustrates how comparative analyses of viral lineages from different hosts can identify positive selection that is otherwise obscured by strong functional constraint. IMPORTANCE: There is a strong interest in correlates of anti-influenza immunity that are protective against diverse virus strains. CD8(+) T cells provide such broad immunity, since they target conserved viral proteins. An important question is whether T-cell immunity is sufficiently strong to drive influenza virus evolution. Although many studies have shown that T cells limit viral replication in animal models and are associated with decreased symptoms in humans, no studies have proven with statistical significance that influenza virus evolves under positive selection to escape T cells. Here we use comparisons of human and swine influenza viruses to rigorously demonstrate that human influenza virus evolves under pressure to fix mutations in the nucleoprotein that promote escape from T cells. We further show that viruses with these mutations have a selective advantage since they are preferentially located on the "trunk" of the phylogenetic tree. Overall, our results show that CD8(+) T cells targeting nucleoprotein play an important role in shaping influenza virus evolution.


Asunto(s)
Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/genética , Subtipo H1N1 del Virus de la Influenza A/inmunología , Gripe Humana/inmunología , Proteínas de Unión al ARN/genética , Selección Genética , Proteínas del Núcleo Viral/genética , Proteínas de la Matriz Viral/inmunología , Animales , Humanos , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H1N2 del Virus de la Influenza A/genética , Subtipo H1N2 del Virus de la Influenza A/inmunología , Subtipo H3N2 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/inmunología , Gripe Humana/virología , Proteínas de la Nucleocápside , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/virología , Porcinos
4.
J Immunol ; 190(3): 1285-96, 2013 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-23275600

RESUMEN

Macrophage dysfunction in obesity and diabetes may predispose to the development of diabetic complications, such as infection and impaired healing after tissue damage. Saturated fatty acids, such as palmitate, are present at elevated concentrations in the plasma of patients with metabolic disease and may contribute to the pathogenesis of diabetes and its sequelae. To examine the effect of lipid excess on macrophage inflammatory function, we determined the influence of palmitate on LPS-mediated responses in peritoneal macrophages. Palmitate and LPS led to a profound synergistic cell death response in both primary and RAW 264.7 macrophages. The cell death had features of apoptosis and necrosis and was not dependent on endoplasmic reticulum stress, ceramide generation, or reactive oxygen species production. Instead, we uncovered a macrophage death pathway that required TLR4 signaling via TRIF but was independent of NF-κB, MAPKs, and IRF3. A significant decrease in macrophage lysosomal content was observed early in the death pathway, with evidence of lysosomal membrane damage occurring later in the death response. Overexpression of the transcription factor TFEB, which induces a lysosomal biogenic program, rescued the lysosomal phenotype and improved viability in palmitate- and LPS-treated cells. Our findings provide new evidence for cross-talk between lipid metabolism and the innate immune response that converges on the lysosome.


Asunto(s)
Proteínas Adaptadoras del Transporte Vesicular/fisiología , Lisosomas/inmunología , Macrófagos Peritoneales/efectos de los fármacos , Palmitatos/toxicidad , Receptor Toll-Like 4/fisiología , Animales , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/fisiología , Muerte Celular/fisiología , Línea Celular Transformada/efectos de los fármacos , Línea Celular Transformada/metabolismo , Línea Celular Transformada/patología , Células Cultivadas/efectos de los fármacos , Células Cultivadas/metabolismo , Células Cultivadas/patología , Complicaciones de la Diabetes/metabolismo , Células HEK293 , Humanos , Inmunidad Innata , Membranas Intracelulares/patología , Metabolismo de los Lípidos/inmunología , Lipopolisacáridos/toxicidad , Lisosomas/patología , Macrófagos Peritoneales/metabolismo , Macrófagos Peritoneales/patología , Ratones , Ratones Endogámicos C57BL , Transducción de Señal/fisiología , Transfección
5.
J Biol Chem ; 288(5): 2923-32, 2013 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-23250746

RESUMEN

Macrophages play a key role in host defense and in tissue repair after injury. Emerging evidence suggests that macrophage dysfunction in states of lipid excess can contribute to the development of insulin resistance and may underlie inflammatory complications of diabetes. Ceramides are sphingolipids that modulate a variety of cellular responses including cell death, autophagy, insulin signaling, and inflammation. In this study we investigated the intersection between TLR4-mediated inflammatory signaling and saturated fatty acids with regard to ceramide generation. Primary macrophages treated with lipopolysaccharide (LPS) did not produce C16 ceramide, whereas palmitate exposure led to a modest increase in this sphingolipid. Strikingly, the combination of LPS and palmitate led to a synergistic increase in C16 ceramide. This response occurred via cross-talk at the level of de novo ceramide synthesis in the ER. The synergistic response required TLR4 signaling via MyD88 and TIR-domain-containing adaptor-inducing interferon beta (TRIF), whereas palmitate-induced ceramide production occurred independent of these inflammatory molecules. This ceramide response augmented IL-1ß and TNFα release, a process that may contribute to the enhanced inflammatory response in metabolic diseases characterized by dyslipidemia.


Asunto(s)
Ceramidas/biosíntesis , Lipopolisacáridos/farmacología , Macrófagos/metabolismo , Palmitatos/farmacología , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Animales , Vías Biosintéticas/efectos de los fármacos , Extractos Celulares , Células Cultivadas , Interleucina-1beta/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/enzimología , Ratones , Ratones Endogámicos C57BL , Factor 88 de Diferenciación Mieloide/metabolismo , FN-kappa B/metabolismo , Esfingolípidos/metabolismo , Receptor Toll-Like 4/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo
6.
Am J Physiol Heart Circ Physiol ; 303(11): H1366-73, 2012 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-23042950

RESUMEN

Diabetes is associated with myocardial lipid accumulation and an increased risk of heart failure. Although cardiac myocyte lipid overload is thought to contribute to the pathogenesis of cardiomyopathy in the setting of diabetes, the mechanism(s) through which this occurs is not well understood. Increasingly, inflammation has been recognized as a key pathogenic feature of lipid excess and diabetes. In this study, we sought to investigate the role of inflammatory activation in the pathogenesis of lipotoxic cardiomyopathy using the α-myosin heavy chain promoter-driven long-chain acylCoA synthetase 1 (MHC-ACS) transgenic mouse model. We found that several inflammatory cytokines were upregulated in the myocardium of MHC-ACS mice before the onset of cardiac dysfunction, and this was accompanied by macrophage infiltration. Depletion of macrophages with liposomal clodrolip reduced the cardiac inflammatory response and improved cardiac function. Thus, in this model of lipotoxic cardiac injury, early induction of inflammation and macrophage recruitment contribute to adverse cardiac remodeling. These findings have implications for our understanding of heart failure in the setting of obesity and diabetes.


Asunto(s)
Cardiomiopatías/patología , Cardiomiopatías/fisiopatología , Corazón/fisiopatología , Lípidos/fisiología , Macrófagos/fisiología , Animales , Cardiomiopatías/metabolismo , Movimiento Celular/fisiología , Coenzima A Ligasas/genética , Coenzima A Ligasas/metabolismo , Citocinas/metabolismo , Modelos Animales de Enfermedad , Metabolismo de los Lípidos/fisiología , Macrófagos/patología , Ratones , Ratones Endogámicos , Ratones Transgénicos , Cadenas Pesadas de Miosina/genética , Cadenas Pesadas de Miosina/metabolismo
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